TiO nanoparticles in seawater: Aggregation and interactions with the green alga Dunaliella tertiolecta.

Titanium dioxide nanoparticles (TiO NPs) have been widely employed in industrial applications, thus rising concern about their impact in the aquatic environment. In this study we investigated the chemical behaviour of TiO NPs in the culture medium and its effect on the green alga Dunaliella tertiolecta, in terms of growth inhibition, oxidative stress, ROS (Reactive Oxygen Species) accumulation and chlorophyll content. In addition, the influence of exopolymeric substances (EPS) excreted by the microalgae on the stability of NPs has been evaluated. The physicochemical characterization showed a high propensity of TiO NPs to form micrometric-sized aggregates within 30min, large enough to partially settle to the bottom of the test vessel. Indeed, an increasing amount of TiO particles settled out with time, but the presence of EPS seemed to mitigate this behaviour in the first 6h of exposure where the main effects in D. tertiolecta were observed. TiO NPs did not inhibit the 72-h growth rate of D. tertiolecta, nor affected the cellular chlorophyll concentration in the range 0.01-10mgL. The time-course of ROS production showed an initial transient increase of ROS in TiO NP-exposed algae compared to the control, concomitant with an enhancement of catalase activity. Interestingly, intracellular ROS was a small fraction of total ROS, the highest amount being extracellular. The occurrence of cell-mediated chemical transformations of TiO NPs in the external medium, related to the presence of EPS, has been evaluated. Our results showed that carbohydrates were the major component of EPS, whereas proteins of medium molecular weight (20-80kDa) were preferentially bound to TiO NPs, likely influencing their biological fate.